Adenine derivatives

ABSTRACT

Novel 2-amino substituted adenine compounds and their use as antiviral compounds are disclosed. In particular the compound 2-amino-9-(2-hydroxyethoxymethyl) adenine is particularly advantageous as an antiviral in that it has high oral absorption and that upon metabolism in mammals, it has the property of being converted to 9-(2-hydroxyethoxymethyl) guanine which is also a highly active antiviral.

This is a division of application Ser. No. 718,105 filed Aug. 27, 1976,abandoned, which is a continuation in part of U.S. Applications Ser. No.662,900 filed Mar. 1, 1976, U.S. Pat. No. 4,199,574, and Ser. No.608,263 filed Aug. 27, 1975, abandoned.

This invention relates to substituted purine compounds and theirpharmaceutically acceptable salts and to methods of preparing them. Inparticular this invention relates to the 9-(2-hydroxyethoxymethyl)derivatives of purines such as adenane, gyanine, thioguanine and2,6-diaminopurine and the pharmaceutically acceptable salts of thesecompounds. In 1971, Schaeffer, et al. (J. Med. Chem., 14, 367 (1971))reported the syntheses of several purine acyclic nucleosides in a studyof adenosine deaminase enzyme-substrate interaction.9-(2-Hydroxyethoxymethyl) adenine in particular was reported and itssubstrate activity with adenosine deaminase measured.

It has now been discovered that substitutes purities of formula (I)##STR1## wherein X is oxygen or sulphur and R¹, R², R³, R⁴, R⁵ and R⁶are various substituents as described herein, have antiviral activityagainst various classes of DNA and RNA viruses both in in vitro and invivo experiments. In particular the compounds are active as anti-viralsagainst cytomegalovirus, adenovirus, in particular adenovirus 5, rhinovirus, Mengo virus and Sindbis virus. They are especially active as ananti-viral against vaccinia, and herpes viruses, including simplex,zoster and varicella, in mammals, which cause such diseases as forexample herpetic keratitis in rabbits and herpetic encephalitis in mice.

According to the present invention there is provided a compound offormula (I) wherein X is sulphur or oxygen, R¹ is hydrogen, halogen,hydroxy, alkoxy, azide, thio, alkylthio, amino, alkylamino ordialkylamino; R² is hydrogen, halogen, alkylthio, acylamino, amino orazide; R³ is hydrogen, straight or branch chain or cyclic alkyl,hydroxyalkyl, benzyloxyalkyl or phenyl; R⁴ is hydrogen, hydroxy oralkyl; R⁵ is hydrogen, hydroxy, amino, alkyl, hydroxyalkyl, benzyloxy,benzoyloxy, benzoyloxymethyl, sulphamoyloxy, phosphate, straight orbranched chain or cyclic acyloxy having from 1 to 8 carbon atoms, orsubstituted carbamoyl group of formula NH.CO--Z wherein Z is alkyl, arylor aralkyl optionally substituted by one or more of sulphonyl, amino,carbamoyl or halogen; R⁶ is hydrogen or alkyl, provided that when X isoxygen and R², R³, R⁴ and R⁶ are hydrogen, R¹ is not amino ormethylamino when R⁵ is hydrogen or hydroxy, or a salt thereof especiallyin the form of a pharmaceutically acceptable salt.

Compounds of formula (I), wherein X is sulphur or oxygen; R¹ ishydrogen, halogen, hydroxy, alkoxy, azide, thio, alkylthio, amino,alkylamino or dialkylamino; R² is hydrogen, halogen, amino or azide; R³is hydrogen, straight or branched chain or cyclic alkyl, hydroxyalkyl,benzyloxyalkyl or phenyl; R⁴ is hydrogen, hydroxy or alkyl; R⁵ ishydrogen, hydroxy, amino, alkyl, hydroxyalkyl, benzoyloxy,benzoyloxymethyl, benzyloxy, sulphamoyloxy, phosphate,carboxypropionyloxy, acetoxy, or substituted carbamoyl group of formulaNH.CO--Z wherein Z is alkyl, aryl or aralkyl optionally substituted byone or more of sulphonyl, amino, carbamoyl, halogen; R⁶ is hydrogen, oralkyl, provided that when X is oxygen and R², R³, R⁴ and R⁶ arehydrogen, R¹ is not amino or methylamino when R⁵ is hydrogen or hydroxy,or a salt thereof, especially in the form of a pharmaceuticallyacceptable salt are preferred.

Cyclic acyloxy is defined to include alicyclic acyloxy and aromaticacyloxy, i.e. aroyloxy such as benzoyloxy.

Especially preferred are compounds of formula (I) as defined abovewherein R⁵ is straight or branched chain acyloxy containing 1 to 5carbon atoms, e.g. formyloxy, acetyloxy, propionyloxy, and pivaloyloxy,or an aroyloxy, e.g. benzoyloxy, and most especially wherein R¹ is aminoor hydroxy, R² is amino, and R³, R⁴, and R⁶ are hydrogen, and theirsalts, especially their pharmaceutically acceptable salts.

Also especially preferred are compounds of formula (I) as defined abovewherein R⁵ is carboxyacyloxy containing 4 to 6 carbon atoms, e.g.carboxypropionyloxy (i.e. 3-carboxypropionyloxy), carboxybutyryloxy(i.e. 4-carboxybutyryloxy) and carboxyvaleryloxy (i.e.5-carboxyvaleryloxy), and most especially wherein R¹ is amino orhydroxy, R² is amino, and R³, R⁴, and R⁶ are hydrogen and most preferredwherein R¹ is hydroxy, R² is amino, and R³, R⁴, and R⁶ are hydrogen, andtheir salts, especially their pharmaceutically acceptable salts.

Compounds of formula (I) as defined above wherein X is oxygen; R¹ ishydrogen, halogen, hydroxy, alkoxy, thio, alkylthio, amino, akylamino,dialkylamino or azide; R² is hydrogen, halogen, amino or azide; R³ ishydrogen, straight or branched chain or cyclic alkyl, hydroxyalkyl orphenyl; R⁴ is hydrogen, or hydroxy; R⁵ is hydrogen, hydroxy, benzoyloxy,hydroxyalkyl, amino, carboxypropionyloxy, acetoxy, benzyloxy,benzoyloxymethyl, phosphate, sulphamoyloxy, substituted carbamoyl groupof formula NH.CO--Z where Z is alkyl, aryl or aralkyl optionallysubstituted by one or more of sulphonyl, amino, carbamoyl, halogen; R⁶is hydrogen, or alkyl, provided that R⁵ is hydroxy only where R¹ isamino, hydroxy, alkylamino, alkylthio, or dialkylamino, and R² is aminoand R⁶ is hydrogen; R⁵ is alkyl-hydroxy only when R¹ is hydroxy; R⁵ ishydrogen only when R¹ is hydroxy or halo; when R⁵ is benzoyloxy R² isnot halogen; R⁵ is acetoxy only when R¹ is hydroxy or amino and R² isamino or both R¹ and R² are halogen; R⁵ is a substituted carbamoyl offormula NH.CO--Z wherein Z is a group CH(NH₂)CH₂ C₆ H₅ only when R¹ isalalkylamino; except that when R⁵ is hydroxy and R is alkylamino then R²is not hydrogen; or a salt thereof, especially in the form of apharmaceutically acceptable salt are particularly preferred.

In particular compounds of formula (I), as hereinbefore defined, where Xis oxygen, R¹ is halogen, amino, hydroxy or alkylthio; R² is amino; R⁵is hydroxy, benzoyloxy, carboxypropionyloxy, acetoxy or hydroxyalkyl andR³, R⁴ and R⁶ are hydrogen, provided that R⁵ is hydroxyalkyl only whenR¹ is hydroxy and R⁵ is acetoxy only when R¹ is hydroxy or amino, aremost preferred and have been found to be highly active. Compounds whereX is sulphur, R¹ is halogen, amino or alkylamino and R², R³, R⁴, R⁵ andR⁶ are hydrogen have also been found to be highly active.

Additionally, the compound where X is oxygen, R¹ is hydroxy, R² isamino, R³, R⁴ and R⁶ are hydrogen and R⁵ is a formyloxy group has beenfound to be highly active.

The preferred halogen substituent is chlorine. As used herein andthroughout the specification the term alkyl is denoted to mean 1 to 12carbon atoms, preferably 1 to 3 carbon atoms.

Salts which are especially convenient for therapeutic use are salts ofpharmaceutically acceptable organic acids such as lactic, acetic, malic,or p-toluenesulfonic acid as well as salts of pharmaceuticallyacceptable mineral acids such as hydrochloric or sulfuric acid.

In a second aspect of the present invention there is provided a methodof preparing a substituted urine of formula (I) or an acid addition saltthereof. ##STR2## wherein X, R¹, R², R³, R⁴, R⁵ and R⁶ are hereinbeforedefined provided that when X is oxygen and R², R³, R⁴ and R⁶ arehydrogen, R¹ is not amino or methylamino when R⁵ is hydrogen or hydroxy;or a salt therof, especially a pharmaceutically acceptable salt thereofcharacterised in that:

(a) where in the compound of formula (I) R⁵ is amino or hydroxy, aprotective group Y is replaced in a compound of formula (II) by an aminoor a hydroxy group; ##STR3##

(b) a compound of formula (III) wherein M and G are precursors of groupsR¹ and R² respectively is converted ##STR4## into a compound of formula(I) where R¹ and R², have the values as defined above;

(c) a compound of formula (IV) ##STR5## is reacted with a compound offormula (V) ##STR6## wherein A is a leaving group and Q is an hydrogenatom or a leaving group;

(d) a ring is closed in a precursor compound having either thepyrimidine or imidazole ring incompletely formed;

(e) a blocking group is removed from a compound of formulation (I)wherein one or both of R¹ and R² is blocked; and where the product ofsaid reaction is a base, optionally converting a compound of formula (I)into an acid addition salt thereof, or where the product is a salt of acompound of formula (I), optionally converting said salt into its baseor another salt thereof.

In the case of method (a) the 9-ether chain is blocked terminally by aprotective group Y which may be an ester, eg an acyloxy group, an amide,eg a phthalimido group, or an arylmethoxy group, eg benzyloxy. In thefirst instance the acyl group may be aliphatic eg acetyl, or aromatic egbenzoyl, both types of acyl group being removed by mild basichydrolysis. In general warming with aqueous methylamine will suffice tobring about the deblocking.

Arylmethoxy blocking groups, such as benzyloxy, are removed byhydrogenolysis, either catalytically, as by hydrogen and Raney nickel orpalladium-on-charcoal, or chemically, as by sodium in liquid ammonia.When sodium in liquid ammonia is used, an excess of ammonia serves assolvent. For catalytic hydrogenolysis an alkanol is the preferredsolvent, although a number of inert, eg non halogen and non-sulphide ormercapto containing solvents may be used provided they dissolve theacyl-blocked substrate, eg such solvents as benzene, tetra hydrofuran ordioxane.

Conversion of a compound of formula (III) ##STR7## into a compound offormula (I), by method (b), can be achieved in numerous different ways.For example one or both of G and M can be converted into a halogen atomby halogenation; into an hydroxy group by hydrolysis; into a thio oralkylthio group by sulphuration; into an hydrogen atom by reduction ordesulphuration; or by formation of the azide. All such methods are wellknown and can be found in "Heterocyclic compounds--Fused PyrimidinesPart II Purines Ed. by D. J. Brown (1971) published Wiley-Interscience".

Alternatively conversion can be brought about by using enzymes, forexample adenosine deaminase efficiently converts a 2,6-diamino compoundto guanine R¹ =OH R² =NH₂, in an aqueous suspension at about 37° C. andinitial pH of about 7.0. Those compounds that contain precursors ofgroups R¹ and R² and can be converted into compounds of formula (I) canbe considered as intermediates in the synthesis of those compounds.

The intermediate of formula (IX) ##STR8## wherein R¹ is a hydroxy oramino group, R² is a hydrogen or halogen atom, or hydrazino group, andR⁵ is a hydroxy or phthalimido group; and of formula (X) ##STR9##wherein R¹ is an NH group and R² is an NH group, are novel compounds andare therefore provided in a further aspect of the invention.

In method (c) the leaving group A is a reactive residue of an organic orinorganic acid, and may therefore be a halogen atom, or a carboxylategroup, and Q is a hydrogen atom or an acyl group. The preferred methodcomprising the condensation of a purine having the desired 2- and6-substitution with an acyl or aralkyl-blocked 2-haloalkoxyethanol forinstance 2-benzoyloxyethoxymethyl chloride, in a strong polar solventsuch as DMF (dimethylsulphoxide) or hexamethylphosphoramide, and in thepresence of a base such as triethylamine or potassium carbonate. Thereaction is preferably carried out at room temperature over an extendedperiod of time ie several days may be required to give reasonableyields.

Alternatively a thermal condensation, eg fusion reaction, may be carriedout to give the product directly. For this reason a suitably substitutedpurine is heated together with an acyloxy-alkoxymethyl carboxylate eg2-oxa-4-butanediol diacetate, in the presence of a catalytic amountstrong acid such as sulphuric acid. Temperatures in excess of 100° C.are generally required, but they should preferably not be greater thanabout 200° C. in order to minimise decomposition. The temperature shouldbe selected such that the mixture of reactants fuse before they undergodecomposition.

The fusion reaction may also be carried out under substantially the samecondition as above, with perhaps somewhat lower temperatures, between a9-acylpurine and the alkoxmethyl carboxylate or halide. Alternativelythe fusion reaction can be carried out using the diester, for instance2-acetoxyethoxymethyl acetate.

Method (d) involves the ring closure of either the imidazole orpyrimidine ring to give the final product. In the case of the imidazolering this may be achieved by reaction of the suitably substitutedprecursor with a one carbon reagent, such as triethylorthoformate, underfor example mildly acidic conditions, at a temperature of about 25° C.and for several hours. A suitable precursor is a substituted pyrimidineof formula (VI) ##STR10## An alternative reagent to use isdiethoxymethyl acetate, when neutral conditions at about 100° C. forabout 10-15 minutes are preferred.

The ring closure of the pyrimidine ring is similar to that for theimidazole ring except that the carbon reagent being added is generallysubstituted, for example by amino. The blocked, substituted carbon isfirst attached to the 2-amino group on a precursor of formula (VII)##STR11## and then deblocked and cyclized. Cyclization may be achievedin some cases by treatment with acid, however this should be done asclose to neutral as possible. In other cases a compound of formula (VII)can be closed by reaction with one of the standard one carbon reagents,under mild conditions. (See J. Org. Chem 81 (1959) p. 6021 A. Yamazakiet al). It is important in this method that the substitution at the 2position, ie R² of formula I, is introduced during ring closure.Suitable substituents in the 6 position, ie R¹, are limited, to suchgroups as thio, amino or hydroxy.

In method (e) substituents R¹ and R² may be blocked by for instancetrimethylsilyl groups. Such a compound will be the product of thecondensation of a trimethylsilylated purine and an ester or diester asin method (c). Such blocking groups are very labile and can be removedby solvolysis with alcoholic or aqueous ammonia, or by alcoholysis.

Alternatively the mercuric chloride salt of a purine can be prepared inthe presence of alkali and then condensed with a haloether in solvent ofthe aromatic organic type. Prior to preparation of the salt however allreactive substituents on the purine must be blocked, and therefore thelast step in this method is the unblocking of the blocked substituents.

In another method, which combines methods (a) and (b) solvolyticdeblocking may be effected at the same time as replacement of a purinering leaving group e.g. halogen, such as by the reaction with liquidammonia. In this case the leaving group or the purine nucleus isreplaced by an amino group at the same as deblocking the side chain, seemethod (a).

In another aspect of the invention there is provided a pharmaceuticalcomposition or preparation comprising a compound of formula (I), whereinX, R, R², R³, R⁴, R⁵ and R⁶ are hereinbefore defined, when R⁶ is alkylgroup it has from 1 to 8 carbon atoms and in all other cases when thesubstituents have an alkyl moiety it has from 1 to 4 carbon atoms; or apharmaceutically acceptable salt thereof; together with apharmaceutically acceptable carrier therefor. In a particular aspect thepharmaceutical composition comprises a compound of formula (I) ineffective unit dosage form.

As used herein the term "effective unit dosage" or "effective unit dose"is denoted to mean a predetermined antiviral amount sufficient to beeffective against the viral organisms in vivo. Pharmaceuticallyacceptable carriers are materials useful for the purpose ofadministering the medicament, and may be solid, liquid or gaseousmaterials, which are otherwise inert and medically acceptable and arecompatible with the active ingredients.

These pharmaceutical compositions may be given parenterally, orally,used as a suppository or pessary, applied topically as an ointment,cream, aerosol, powder, or given as eye or nose drops etc., depending onwhether the preparation is used to treat internal or external viralinfections.

For internal infections the compositions are administered orally orparenterally at dose levels, calculated as the free base, of about 0.1to 250 mg per kg, preferably 1.0 to 50 mg per kg, of mammal body weight,and are used in man in a unit dosage form, administered, eg a few timesdaily, in the amount of 1 to 250 mg per unit dose.

For oral administration, fine powders or granules may contain diluting,dispersing and/or surface active agents, and may be presented in adraught, in water or in a syrup; in capsules or sachets in the dry stateor an a non-aqueous solution or suspension, wherein suspending agentsmay be included; in tablets, wherein binders and lubricants may beincluded; or in a suspension in water or a syrup. Where desirable ornecessary, flavouring, preserving, suspending, thickening or emulsifyingagents may be included. Tablets and granules are preferred, and thesemay be coated.

For parenteral administration or for administration as drops, as for dyeinfections, the compounds may be presented in aqueous solution in aconcentration of from about 0.1 to 10% more preferably 0.1 to 7%, mostpreferably 0.2% w/v. The solution may contain antioxidants, buffers,etc.

Alternatively for infections of the eye, or other external tissues e.g.mouth and skin the compositions are preferably applied to the infectedpart of the body of the patient as a topical ointment or cream. Thecompounds may be presented in an ointment, for instance with a watersoluble ointment base, or in a cream, for instance with an oil in watercream base, in a concentration of from about 0.1 to 10%, preferably 0.1to 7%, most preferably 1% w/v.

Of the compounds of formula (I), 9-(2-hydroxyethoxymethyl)guanine (R¹=OH, R² =NH₂), 2-amino-9-(2-hydroxyethoxymethyl)adenine (R¹ =R² =NH₂)and their esters are most preferred, particularly because of theirextremely high antiviral activity against herpes viruses. Additionally 2amino-6-chloro-9{(2-benzoyloxyethoxy)methyl)purine,9(2-benzoyloxyethoxymethyl)guanine, 9-(3-hydroxypropoxymethyl)guanine, 2amino-6-methylthio-9-(2-hydroxyethoxymethyl)purine,9-{2-(3-carboxypropionyloxy)ethoxymethyl)guanine,9-(2-acetoxyethoxymethyl)-2,6-diamino purine, 6-chloro-9-ethylthiomethylpurine, 9-ethylthiomethyladenine, 9-ethylthiomethyl-6-methylamino-purinealso show high activity against herpes viruses and vaccinia.

In yet a further aspect of the invention there is provided a method oftreating viral infections in mammals which comprises the administrationof an effective non-toxic anti-viral amount, as hereinbefore defined ofa substituted purine of formula (I), or a pharmaceutically acceptablesalt thereof. Administration is preferably by topical application or bythe oral or parenteral route.

The invention will now be illustrated with reference to the followingexamples.

EXAMPLE 1 6-Chloro-9-(2-benzoyloxyethoxymethyl)purine

A solution of benzonitrile (103 g) in ethylene glycol (310 g) was heatedat reflux under substantially anhydrous conditions for 3 days. Thereaction mixture was cooled and added to a mixture of ice and water(about 300 ml). The resulting mixture was extracted with ether (3×300ml) and the combined ether extract back-washed with water (2×300 ml) andthen with a saturated sodium chloride solution (300 ml). The ethersolution was dried over anhydrous sodium sulfate. The ether wasevaporated and the residual oil distilled to give 108 g (65% oftheoretical) of ethylene glycol monobenzoate, b.p. 132°-135° C./1.5 mmHg.

A cold (0° C.) mixture of ethylene glycol monobenzoate (166 g) andparaformaldehyde (30 g) in dry dichloroethane was saturated with dry HClwith stirring for 3 hours. The pinkish red liquid was dried over calciumchloride and the volatile components removed on a rotary evaporator at30° C. to give 1-benzoyloxy-2-chloromethoxyethane (215 g). The Infra-Redspectrum indicated the absence of a hydroxyl group.

Following substantially the procedure of Schaeffer, et al. supra butwith minor modifications, 1-benzoyloxy-2-chloromethoxyethane (4.4 g) wasadded with stirring to a dry solution of 6-chloropurine (3.1 g) andtriethylamine (6.5 ml) in dimethylformamide (50 ml). The reaction wasexothermic, immediately precipitating, triethylamine hydrochloride. Thereaction mixture was stirred at room temperature for 24 hours andfiltered. The filtrate was evaporated on a rotary evaporator at 70° C.The remaining thick amber oil was dissolved in the minimum amount ofbenzene and purified by column chromatography using a silica gel column.Benzene elution removed a trace of an unidentified material. Elutionwith ether first remove a small amount of ethylene glycol monobenzoatethen 6-chloro-9-(2-benzoyloxyethoxymethyl)purine. Recrystallization fromether gave a white material, m.p. 108.5°-111° C.

EXAMPLE 2 9-(2-Hydroxyethoxymethyl)adenine (I; R¹ =NH₂, R² =H)

A solution of 6-chloro-9-(2-benzoyloxyethoxymethyl)purine (50 g) andammonia (31 g) in methanol (120 ml) was heated in a bomb at 95° C. for18 hours. The reaction mixture was removed from the bomb and the solventevaporated under reduced pressure at 50° C. The resulting solid wastriturated first with water and then, after drying, with chloroform. Thesolid, 9-(2-hydroxyethoxymethyl)adenine, was retained, and the aqueoustriturate was extracted several times with chloroform. The aqueousextracts were treated with a strongly basic ion exchange resin and thenevaporated to dryness under reduced pressure. The remaining solid wascombined with that from the trituration step and recrystallized fromisopropanol to give 22 g (70% of theoretical) of white9-(2-hydroxyethoxymethyl)adenine, m.p. 199.5°-200° C.

EXAMPLE 3 2-Chloro-9-(2-hydroxyethoxymethyl)adenine

Following the procedure of Example 1, 2,6-dichloropurine was condensedwith 1-benzoyloxy-2-chloromethoxyethane to give a 41% yield of2,6-dichloro-9-(2-benzoyloxy ethoxymethyl)purine, m.p. 121°-125° C. Thiswas treated according to the procedure of Example 2 to give simultaneousammonolysis of the 6-chloro and benzoyloxy groups yielding2-chloro-9-(hydroxyethoxymethyl)adenine, in 94% yield, m.p. 188°-190° C.after recrystallization from isopropanol.

EXAMPLE 4 2-Amino-9-(2-hydroxyethoxymethyl)adenine I; R¹ =R² =NH₂

2-Chloro-9-(2-hydroxyethoxymethyl)adenine (0.83 g) was added withstirring to 95% hydrazine (11 ml) and the resulting solution stirred 1hour under nitrogen. Excess hydrazine was removed by distillation in arotary evaporator at a bath temperature of 30° C. followed byco-evaporation twice with ethyl acetate. The residue was thoroughlytriturated with methanol to give a quantitative yield (0.8 g) of whiteto cream colored-2-hydrazine-9-(2-hydroxyethoxymethyl)adenine, m.p.220°-222° C.

A solution of this hydrazine derivative (0.85 g) in 5% aqueous aceticacid (30 ml) was chilled to 5°-8° C., and a cold (0° C.) solution ofsodium nitrite (0.3 g) in water (65 ml) was added in one portion. Thereaction mixture was stirred in an ice bath for one hour and thenfiltered, washed with cold water and dried at room temperature to give2-azido-9-(2-hydroxyethoxymethyl)adenine (0.86 g). Recrystallizationfrom water gave a light mauve product, m.p. 191°-192° C.

A mixture of 2-azido-9-(2-hydroxyethoxymethyl)adenine (0.41 g) and 10%palladium on charcoal (80 mg) in ethanol (41 ml) was shaken underhydrogen at 50 psi for 3.5 hours. The catalyst was removed by filtrationthrough a celite pad and washed thoroughly with ethanol and water. Thesolvent was removed by evaporation under reduced pressure giving aquantitative yield (0.37 g) of nearly pure (one spot on TLC)2-amino-9-(2-hydroxyethoxymethyl)adenine, m.p. 183°-184° C. afterrecrystallization from n-propanol.

EXAMPLE 5 9-(2-Hydroxyethoxymethyl)guanine I; R¹ =OH, R² =NH₂)

Solid sodium nitrite (0.97 g) was added at room temperature withstirring over a period of one hour to a solution of2-chloro-9-(2-hydroxyethoxymethyl)adenine (0.5 g) in glacial acetic acid(10 ml). The reaction mixture was stirred for an additional 41/2 hours.The white solid was removed by filtration, washed with cold acetic acidand then well triturated with cold water to remove the sodium acetatepresent. The solid product was retained. The combined acetic acidfiltrate and wash was evaporated at reduced pressure and 40° C. bathtemperature and the residual oil triturated with cold water. Theresulting solid material was combined with the previously isolated solidand the combined solids dried and recrystallized from ethanol to give2-chloro-9-(2-hydroxyethoxymethyl)hypoxanthine (0.25 g), m.p.>310° C.Elemental analysis and NMR spectrum were consistent with this structure.

A mixture of 2-chloro-9-(2-hydroxyethoxymethyl)hypoxanthine (0.375 g)and methanol (80 ml) saturated with anhydrous ammonia was heated in abomb at 125° C. for 5 hours. The bomb was cooled in an ice bath and thereaction mixture removed. Solvent and excess ammonia were removed underreduced pressure at 50° C. After the residue was triturated with coldwater to remove the ammonium chloride formed, the remaining solid wasdried and then recrystallized from methanol to give pure9-(2-hydroxyethoxymethyl)guanine (0.24 g), m.p. 256.5°-257° C.

EXAMPLE 6 9-(2-Hydroxyethoxymethyl)guanine (I; R¹ =OH, R² =NH₂)

A mixture of guanine (2.0 g), ammonium sulfate (1.5 g), andhexamethyldisilazane (126 g) was heated at reflux under nitrogenovernight. Excess hexamethyldisilazane was removed by distillation atreduced pressure. Dry benzene (10 ml) was added to the residual oil andany remaining ammonium sulfate removed by filtration. To this solutionwas added triethylamine (4 ml) and a solution of2-benzoyloxyethoxymethyl chloride (2.8 g) in dry benzene (15 ml) and themixture heated at reflux under nitrogen overnight. The solvent wasevaporated in a rotary evaporator at reduced pressure and the residuedissolved in 95% ethanol. The solution was warmed on a steam bath for1/2 hour to effect hydrolysis of the silyl groups. The ethanol was thenevaporated and the residual solid thoroughly washed with water, filteredand dried. Recrystallization from methanol and then from water (residualguanine was insoluble in the hot solvents and was removed by filtration)afforded 9-(2-benzoyloxyethoxymethyl)guanine (0.58 g., 14% oftheoretical), m.p. 222°-226° C. A later condensation of thetris-(trimethylsilyl)guanine with a 60% excess of2-benzoyloxyethoxymethyl chloride gave a 32% yield of9-(2-benzoyloxyethoxymethyl)guanine.

9-(Benzoyloxyethoxymethyl)guanine (0.58 g) and methanol (80 ml)saturated with ammonia were heated in a bomb at 80° C. for 16 hours. Thereaction mixture was removed from the bomb and the solvent evaporatedunder reduced pressure. The residue was thoroughly washed with ether andthen recrystallized from methanol to give9-(2-hydroxyethoxymethyl)guanine (0.31 g., 75% of theoretical), m.p.250.5°-257° C.

EXAMPLE 7 2-Amino-9-(2-hydroxyethoxymethyl)adenine (I; R¹ R² =NH₂)

A mixture of 2-chloro-9-(2-hydroxyethoxymethyl)adenine (4.4 g) andliquid ammonia (40 ml) was heated at 120° C. in a bomb for 19 hours. Thereaction mixture was removed from the bomb and excess ammonia removed invacuo. The residue was partitioned between ether and water and theaqueous layer washed twice more with ether and once with chloroform. Theaqueous solution was then treated with an excess of a strongly basic ionexchange resin to convert the ammonium chloride to ammonia andevaporated to dryness in vacuo. The residue was dissolved in a minimalamount of ethanol, Florisil (Registered Trade Mark) (10 g) added, andthe mixture evaporated in dryness. The residue was transferred to aprepared column of Florisil in chloroform and eluted with 5%methanol/95% chloroform (v/v). Fractions (30 ml) were collected andexamined by TLC (silica gel plates developed in 10% methanol/90%chloroform) for the presence of product. The first 40 fractionscontained starting material but no product and were discarded.Succeeding fractions which were found to contain the desired productwere combined and evaporated to dryness. The residue was recrystallizedfrom ethanol (with activated charcoal treatment) and from n-propanol togive 2-amino-9-(2-hydroxyethoxymethyl) adenine (0.5 g, 12% oftheoretical), m.p. 183°-184° C.

Florisil is an activated magnesium silicate widely used in columnchromatography.

EXAMPLE 8 2-Amino-6-chloro-9-(2-benzoyloxyethoxymethyl)purine

A suspension of potassium carbonate (4.1 g) and 2-amino-6-chloropurine(5.0 g) in dry dimethylformamide (120 ml) was stirred for 20 minutes.2-Benzoyloxyethoxymethyl-chloride (6.3 g) was added and the resultingpink suspension stirred at room temperature for 6 days. The reactionmixture was then poured into a mixture of ice and water (210 ml) withvigorous stirring. The aqueous mixture was extracted with chloroform(3×200 ml), and the chloroform solution obtained was washed with a 5%acetic acid solution and then twice with water. The chloroform solutionwas dried over anhydrous sodium sulfate and evaporated in vacuo. Theresidue was redissolved in a minimal amount of chloroform and theresulting solution applied to a column containing silica gel (200 g) inchloroform. The column was eluted with 2% methanol/98% chloroform andthe eluant collected in 30 ml cuts. The fractions containing2-amino-6-chloro-9-(2-benzoyloxyethoxymethyl)purine (determined by TLC)were combined, evaporated and the residue recrystallized from benzene togive 2-amino-6-chloro-9-(2-benzoyloxyethoxymethyl)purine (2.0 g., m.p.130°-133.5° C.).

EXAMPLE 9 9-(2-Hydroxyethoxymethyl)thioguanine (I; R¹ =SH, R² =NH₂)

Thiourea (0.28 g) was added to a refluxing solution of2-amino-6-chloro-9-(2-benzoyloxyethoxymethyl)purine (1.27 g) inisopropanol (40 ml). The reaction mixture was heated at reflux for 11/2hours and then chilled. 9-(2-benzoyloxyethoxymethyl)thioguanine (0.58 g,m.p. 199°-202° C.) was removed by filtration. This was added to 40 ml ofaqueous ammonia (about 7 N) and the mixture heated on a steam bath for10 minutes and then stirred overnight at room temperature. The water andammonia were removed in vacuo and the residue triturated with acetoneand then with ether to remove benzamide. The remaining solid wasrecrystallized twice from water to give yellow flakes of9-(2-hydroxyethoxymethyl)thioguanine (0.26 g, m.p. 251°-254° C.).

EXAMPLE 10 9-(2-Hydroxyethoxymethyl)guanine (I; R¹ =OH, R² =NH₂)

To a solution of 2-amino-9-(2-hydroxyethoxymethyl)adenine (0.22 g) inwater (30 ml) was added a suspension of adenosine deaminase in aqueousammonium sulfate (0.44 ml, containing 4.4 mg of the enzyme). Thereaction mixture, which initially had a pH of 7.0, was heated at 37° C.for 18 hours, at which time the pH was 8.5 and TLC (silica gel platesdeveloped in 15% methanol-85% chloroform) indicated a single productdifferent from starting material. The reaction mixture was thoroughlychilled in an ice bath and the resulting white solid removed byfiltration and thoroughly washed with cold water. The product was driedat 100° C./0.1 mm Hg for 16 hours to give 0.20 g of9-(2-hydroxyethoxymethyl)guanine. 1/4 H₂ O, the structure of which wasconfirmed by m.p., TLC, U.V., NMR and mass spectroscopy analysis.Recrystallization from methanol gave the anhydrous9-(2-hydroxyethoxymethyl)guanine.

EXAMPLE 11 2-Amino-9-(2-hydroxyethoxymethyl)adenine hydrochloride

2-Amino-9-(2-hydroxyethoxymethyl)adenine (0.25 g) was dissolved in hotethanol (50 ml) and the solution cooled in an ice bath. To the coolsolution was added sufficient HCl-saturated ethanol to give a pH of 1.0.Dry ether (50 ml) was then added and the mixture thoroughly chilled. Theresulting flesh-colored solid was removed by filtration and washed withcold ethanol. Recrystallization from methanol yielded2-amino-9-(2-hydroxyethoxymethyl)adenine hydrochloride (0.21 g., 74% oftheoretical), m.p. 207°-211° C. Elemental analysis and NMR support thisstructure.

EXAMPLE 12 Properties of 9-(2-hydroxyethoxymethyl)guanine)

9-(2-Hydroxyethoxymethyl)guanine was determined by U.V. to be soluble ineither water or 0.1 N aqueous hydrochloric acid to the extent of about0.2%. It was stable in either 0.01 N or 0.1 N aqueous hydrochloric acidat 26° C. and at 37° C., showing no indication of hydrolysis after oneweek. However, in 1 N aqueous hydrochloric acid at either 26° C. or 37°C., it slowly hydrolyzed to guanine within one week. 13.

Preparation of 2-amino-9-(2-hydroxyethoxymethyl)adenine

2,6-dichloro-9-(2-benzoyloxyethoxymethyl)purine 10 g, sodium azide 3.5 gand 54 ml. of 1:1 (v/v) ethanol-water was refluxed with stirring for31/2 hours at 110°-120° C. TLC of reaction mixture shows completereaction. On overnight cooling the oil solidified and was filtered,washed with ethanol and water and recrystallized from ethanol.Quantitative yield m.p. 124°-125° C. Compound is labile in aqueous baseand sensitive to sunlight.

1.2 g of 2,6-diazido-9-(2-benzoyloxyethoxymethyl)purine was dissolved in150 ml. 1:1 methanol tetrahydrofuran and the solvent was put in Paarhydrogenation apparatus containing 33 mg. of 10% palladium charcoalwhich had been wetted with water and shaken under 50 psi hydrogen for4.5 hours. The catalyst was filtered and washed well with water andmethanol TLC showed complete reduction. Solution was evaporated todryness giving a yield of 87% of 2,6-diamino intermediate.

The residue was dissolved in 40% aqueous methylamine minimum amount andwas heated for 1/2 hour on a steam bath, cooled and extracted 3 timeswith an equal volume of ether. The ether extracts were back-washed withwater and the combined aqueous phase evaporated to dryness. The residuewas recrystallised from ethanol to give 89% yield of the compound2-amino-9-(2-hydroxyethoxymethyl)adenine m.p. 182°-183.5° C. 14.

Preparation of 2,6-Dichloro-9-(2-acetyloxyethoxymethyl)purine

2,6-Dichloropurine 5.5 g and 2-oxa-1,4-butanediol diacetate 513 g wereplaced in a flask and partially evacuated and then heated to 138° C.,initially the mixture was too thick to stir but gradually gave rise to amelt which was stirred, and after 20 minutes was completely melted andthe reaction mixture heated for 10 minutes more (total heat time=30minutes). The mixture was then cooled to room temperature andpara-toluenesulphonic acid 150 mg was added, the vacuum reapplied andheating resumed with stirring. After 20 minutes heating vigorousbubbling was noted; the mixture was cooled to room temperature;chloroform was added, and the solution was extracted once with saturatedaqueous sodium bicarbonate and once with water. The chloroform phase wasdried over anhydrous sodium sulphate and evaporated. The residual oilwas dissolved in benzene and applied to a column of 200 g. of silica gelin benzene. Benzene elution removed the acetate by-products. Ethersolution gave 2,6-dichloro-9-(2-acetyloxyethoxymethyl)purine (64%yield). Recrystallization from benzene gave lustrous white flakes, m.p.96°-99° C.

EXAMPLE 15 2,6-Diamino-9-(2-benzoyloxyethoxymethyl)purine

A mixture of 2,6-diaminopurine monohydrate (2.0 g), ammonium sulfate(1.32 g), and hexamethyldisilizane (100 g) was heated at reflux undernitrogen for 18 hours. The solvent was evaporated under reduced pressureand the residual oil dissolved in a minimal amount of benzene. To thebenzene solution was added 2-benzoyloxyethoxymethyl chloride (2.56 g),triethylamine (2 ml) and benzene (55 ml). This reaction mixture washeated at reflux under nitrogen for 18 hours. Additional2-benzoyloxyethoxymethyl chloride (2.56 g) and triethylamine (2 ml) wereadded, and heating at reflux was continued for an additional 6 hours.The solvent was evaporated under reduced pressure and the residuedigested on a steam bath for 30 minutes in 95% ethanol (40 ml). Thesolvent was evaporated and the gummy residue recrystallized fromethanol, twice from methanol and finally from water to give2,6-diamino-9-(2-benzoyloxyethoxymethyl)purine as a yellow solid, m.p.205° C., in 7.5% yield.

EXAMPLE 16 2-Amino-6-benzyloxy-9-(2-hydroxyethoxymethyl)purine

A solution of sodium benzylate in benzyl alcohol was prepared fromsodium (2.58 g) and benzyl alcohol (28 ml). The solution was heated to120° C. and 2-amino-6-chloro-9-(2-benzoyloxyethoxymethyl)purine (3.47 g)was added over a 10 minute period. The reaction mixture was headedovernight with stirring at 120°-130° C. and then poured into a mixtureof ice and water. The resulting mixture was extracted thoroughly withchloroform. The aqueous phase was neutralized with acetic acid, giving awhite precipitate which was recrystallized from methanol and then fromwater to give analytically pure2-amino-6-benzyloxy-9-(2-hydroxyethoxymethyl)purine, m.p. 291°-292° C.(dec.) in 50% yield.

This compound may be reductively cleaved, e.g. with hydrogen,palladium/charcoal in methanol, to give9-(2-hydroxyethoxymethyl)guanine.

EXAMPLE 17 9-(2-Hydroxyethoxymethyl)guanine

2-Mercaptoethanol (0.75 ml) dissolved in 1 M methanolic sodium methylate(7.5 ml) was added to2-amino-6-chloro-9-(2-benzoyloxyethoxymethyl)purine (0.89 g) in methanol(150 ml). The reaction mixture was heated at reflux for 3 hours undernitrogen. The solvent was evaporated under reduced pressure and theresidue dissolved in water. The aqueous solution was heated on a steambath for 2 hours, chilled, and acidified to pH 5.0 with acetic acid. Theresulting white solid was removed by filtration, washed well with icecold water and ether, and then recrystallized from methanol to give a45% yield of 9-(2-hydroxymethoxymethyl)guanine.

EXAMPLE 18 9-Ethylthiomethyladenine

A reaction mixture containing chloromethyl ethyl sulfide (5.53 g),6-chloropurine (7.73 g) and triethylamine (5.57) in dimethylformamide(50 ml) was allowed to stand at room temperature for three days. It wasfiltered and the filtrate evaporated. The semi-solid residue wasdissolved in chloroform (ca. 80 ml), placed on a column of Florisil(activated magnesium silicate) (360 g) in chloroform, and eluted withchloroform. The initial eluate (ca. 500 ml) was discarded; the next 2.8liters were collected and evaporated. Ligroin (ca. 50 ml) was added tothe residual oil, the mixture chilled, and the resulting off whitecrystals of 6-chloro-9-ethylthiomethylpurine collected by filtration andwashed with ligroin; yield 3.3 g (m.p. 78°-81° C). Recrystallizationfrom ether-ligroin gave white needles, m.p. 81°-82.5° C.

6-Chloro-9-ethylthiomethylpurine (1.5 g) and liquid ammonia (20 ml) wereplaced in a bomb and heated at 60° C. overnight. Evaporation of theammonia gave a residue which was treated with cold water, filtered, andwashed with cold water. This yielded 9-ethylthiomethyladenine (1.17 g)as a white solid, m.p. 140°-142° C. Recrystallization from ethanol gavewhite plates, m.p. 142°-143° C.

EXAMPLE 19 9-(2-Hydroxyethylthiomethyl)adenine

2-Acetoxyethanethiol was prepared according to the method of Miles andOwen, J. Chem. Soc., 817 (1952). Acetic anhydride (102.1 g) was addedover 1.25 hours to a mixture of 2-mercaptoethanol (78.1 g) and a 10%solution of sulfuric acid in acetic acid (3 ml). External cooling wasused during the addition to keep the reaction temperature below about40° C. After the addition of acetic anhydride was complete, the reactionmixture was heated at 65° C. for one hour and allowed to stand at roomtemperature overnight. Ether (500 ml) was added to the reaction mixtureand the resulting solution washed with water (3×100 ml) and with brine(1×100 ml). The ether solution was dried over anhydrous sodium sulfateand sodium bicarbrate. Distillation gave 2-acetoxyethanethiol (77.5 g,bp 57°-67° C. at 10 mm Hg).

Hydrogen chloride gas was introduced into a mixture of2-acetoxyethanethiol (24.0 g) and paraformaldehyde (6.0 g) at a moderaterate with external cooling for 3 hours. Calcium chloride (25 g) wasadded and the reaction mixture allowed to stand in a salt-ice bath for 4hours. Methylene chloride (200 ml) was added, the reaction mixturefiltered and the solvent evaporated with rigorous exclusion of moisture.The residual oil was distilled to give 2-acetyloxyethyl chloromethylsulfide (18.5 g, bp 82°-87° C. at 3 mm Hg).

Following the procedure of Schaeffer, et al. J. Med. Chem., 14, 367(1971), 2-acetoxyethyl chloromethyl sulfide (3.37 g) was added to amixture of 6-chloropurine (3.09 g), triethylamine (2.23 g) anddimethylformamide (20 ml). The reaction mixture was stirred at roomtemperature for 90 hours and the solvent then removed under reducedpressure. The residual oil was dissolved in chloroform (30 ml) andplaced on a column of Florisil (200 g) in chloroform. Elution withchloroform resulted in the desired product in 2.1 liters of eluate(after discarding the initial 300 ml collected). The solvent wasevaporated to give a residual oil. Ether-pet.ether was added and themixture chilled to give 9-(2-acetoxyethylthiomethyl)-6-chloropurine (1.4g, mp 82°-87° C.). Recrystallization from ether gave white prisms m.p.89°-91° C.).

9-(2-Acetoxyethylthiomethyl)-6-chloropurine (1.1 g) and ammonia (20 ml)were placed in a bomb and heated at 60° C. for 24 hours. The reactionmixture was removed from the bomb and the ammonia allowed to evaporate.The residue was triturated with cold water, filtered and washed withadditional cold water. The resulting crude9-(2-hydroxyethylthiomethyl)adenine (0.73 g) melted at 166°-169.5° C.Recrystallization from ethanol gave white plates, m.p. 170°-172° C.

EXAMPLE 20 9-(2-(3-Carboxypropionyloxy)ethoxymethyl)guanine

A mixture of 9-(2-hydroxyethoxymethyl)guanine (0.25 g), succinicanhydride (0.55 g) and pyridine (50 ml) was heated under anhydrousconditions on a steam bath overnight. The solvent was evaporated underreduced pressure at <40° C., the last trace being removed azeotropicallywith toluene. The residue was triturated with acetone and the productremoved by filtration. Recrystallization from methanol afforded9-(2-(3-carboxypropionyloxy)ethoxymethyl)guanine, m.p. 203°-207° C.(sinter 190° C.), in 44% yield.

EXAMPLE 21 9-Ethylthiomethyl-N-6-methyladenine

6-Chloro-9-ethylthiomethylpurine (1.2 g) and methylamine (20 ml) wereplaced in a bomb and heated at 60° C. for 24 hours. The excessmethylamine was then allowed to evaporate at room temperature. Theresidue was triturated with ice water (10 ml) and filtered.Recrystallization of the tan solid from ether with charcoal (Darco (O)treatment gave 9-ethylthiomethyl-N-6-methyladenine (0.64 g), m.p.111.5°-113° C.

EXAMPLE 22 9-(2-Aminoethyoxymethyl)adenine

To a stirred, ice-bath cooled dispersion of sodium hydride (4.0 g of a60% dispersion in mineral oil) in dimethylformamide (500 ml) was addedadenine (13.5 g). After one hour, the ice-bath was removed and themixture stirred at ambient temperature for an additional 3 hours. Asolution of N-(2-chloromethoxyethyl)phthalimide (23.9 g) indimethylformamide (100 ml) was added dropwise over 0.5 hour. Thereaction mixture was stirred at ambient temperature for 18 hours andthen poured with stirring over ice water (2 liters). The resulting solidwas recrystallized from 2-methoxyethanol and then from dimethylformamideto give 9-(2-phthalimidoethoxymethyl)adenine (15.0 g), m.p. 256°-258° C.

A mixture of 9-(2-phthalimidoethoxymethyl)adenine (3.38 g), hydrazinehydrate (1.0 ml of an 85% aqueous solution) and ethanol (150 ml) washeated at reflux for 2 hours. 2-Methoxyethanol (50 ml) was added andrefluxing continued for an additional 2 hours. The solvent was removedunder reduced pressure and 1 N hydrochloric acid (50 ml) added. Themixture was stirred at ambient temperature for 30 minutes and allowed tostand at 0° C. overnight. The solution was filtered and the filtratediluted with water (100 ml) and stirred with excess basic ion exchangeresin {Rexyn 201(OH)} until a negative silver nitrate test was obtained.It was then filtered and most of the water removed under reducedpressure. The aqueous solution was cooled and the resulting solidremoved by filtration. This was recrystallized from isopropanol to give9-(2-aminoethoxymethyl)adenine (1.0 g), m.p. 170°-171° C.

EXAMPLE 23 2,6-Diamino-9-(2-acetoxyethoxymethyl)purine

Following the procedure of Example 13,2,6-dichloro-9-(2-acetyloxyethoxymethyl)purine was converted to2,6-diazido-9-(2-acetyloxyethoxymethyl)purine, m.p. 69°-70° C. (fromethanol water), in 83% yield. And this was catalytically reduced byhydrogen over 10% palladium on charcoal in methanol at 50 psi to give2,6-diamino-9-(2-acetoxethoxymethyl)purine, m.p. 158°-159° C. (fromethanol), in 88% yield.

EXAMPLE 24 2-Amino-6-methylthio-9-(2-hydroxyethoxymethyl)purine

A mixture 9-(2-Hydroxyethoxymethyl)thioguanine (0.7 g), methyl iodide(0.2 ml), Dowex HCO₃ ⁻ resin (1.7 g) and methanol (200 ml) was stirredovernight at room temperature. The resin was removed by filtration,washed with methanol and the combined filtration evaporated, and theresidue recrystallized from isopropanol and then from acetone to give2-amino-6-methylthio-9-(2-hydroxyethoxymethyl)purine, m.p. 188°-191° C.in 38% yield.

EXAMPLE 25 9-(3-Hydroxypropoxymethyl)guanine

Sodium benzoate (96.32 g) in DMF (690 ml) was heated to 80° C., and1-chloro-3-hydroxypropane (63.06 g) was added over 15 minutes. Thetemperature increased to 135° C., and the reaction mixture was heated at135°-175° C. for 3 hours. Filtration removed 38 g of sodium chloride(97% of theory). The filtration was partially evaporated at reducedpressure at <40° C. The concentrated filtrate was poured into ice waterand extracted well with ether. The combined ether extracts were washedwith water, dried over anhydrous sodium sulfate, and evaporated. Theresidual oil was distilled through a Vigreux column to give3-benzoyloxy-1-propanol (85.2 g), b.p. 124°-132° C. at 0.055 mm Hg.

Anhydrous hydrogen chloride was bubbled into a solution of3-benzoyloxy-1-propanol (15.02 g) and paraformaldehyde (2.49 g indichloromethane (35 ml) for 1 hour at 0° C. The solvent was evaporatedunder reduced pressure at <40° C. giving a 92% yield of crude3-benzoyloxypropoxymethyl chloride which was used without purification.

A solution of trimethylsilylated guanine in benzene (25 ml) prepared asin Example 6 (from 2.0 g of guanine) containing triethylamine was heatedat reflux and 3-benzoyloxypropoxymethyl chloride (2.96 g) dissolved inbenzene (15 ml) was added over a 3 hour period. The reaction mixture washeated at reflux under nitrogen overnight. The solvent was removed underreduced pressure, and 95% ethanol and methanol were added to theresidual oil. The mixture was heated on a steam bath for several minutesand the solvent then evaporated under reduced pressure. Chloroform (200ml) was added and the resulting solid removed by filtration. The solidwas dissolved in a minimal amount of DMF, filtered (to remove anyguanine present) and reprecipitated by the addition of water.Recrystallization from methanol (with charcoaling) gave9-(3-benzoyloxypropoxymethyl)guanine (0.94 g) as a pale yellow solid,m.p. 198°-201° C.

A mixture of 9-(3-benzoyloxypropoxymethyl)guanine (0.5 g) and aqueous45% methylamine (10 ml) was stirred overnight at room temperature.Excess methylamine and water was evaporated at <30° C. under reducedpressure and the residue recrystallized from ethanol to give9-(3-hydroxypropoxymethyl)guanine (0.24 g), m.p. 223° C. (withresolidification), as the 1/2 hydrate.

EXAMPLE 26 Oil in Water Cream base

    ______________________________________                                        9-(2-hydroxyethoxymethyl)guanine                                                                       5.0 g                                                Lanolin, Anhydrous       20.0 g                                               Polysorbate 60           4.0 g                                                Sorbitan Monopalmitate   2.0 g                                                Light Liquid Paraffin    4.0 g                                                Propylene Glycol         5.0 g                                                Methyl Hydroxybenzoate   0.1 g                                                Purified Water           to 100.0 g                                           ______________________________________                                    

EXAMPLE 27 Water Soluble Ointment Base

    ______________________________________                                        2-amino-9-(2-hydroxyethoxymethyl)adenine                                                                 0.3 g                                              Glycerol                   15.0 g                                             Macrogol 300               20.0 g                                             Polyethylene Glycol 1500   64.5 g                                             ______________________________________                                    

EXAMPLE 28 Tablet--(Total weight 359 mg)

    ______________________________________                                        9-(2-hydroxyethoxymethyl)guanine                                                                        100 mg                                              Lactose                   200 mg                                              Starch                    50 mg                                               Polyvinylpyrrolidone      5 mg                                                Magnesium Stearate        4 mg                                                ______________________________________                                    

EXAMPLE 29

A solution of 9-(2-hydroxyethoxymethyl)guanine (4.73 g) in 97% formicacid (24 ml) was stirred at room temperature overnight. The ambersolution was diluted with about 200 ml of dry ether and chilled. Theresulting white precipitate was filtered, dried and recrystallized fromdry dimethylformamide to give 9-(2-formyloxyethoxymethyl)guanine (3.6 g,m.p. 225°-227° C.).

EXAMPLE 30

A solution of 2-amino-9-(2-hydroxyethoxymethyl)adenine (0.5 g) in 97%formic acid (2.5 ml) was stirred in an ice bath for 3 hours, and then atroom temperature overnight. Dry ether (80 ml) was added and the mixturechilled. The solid was removed by filtration and dissolved in hotacetonitrile (125 ml); residual solids were removed by filtration. Thefiltrate was applied to a column containing silica gel (14 g) inacetonitrile. The column was eluted with dry acetone. The eluate wasevaporated and the residual solid recrystallized from acetonitrile togive 2-amino-9-(2-formyloxyethoxymethyl)adenine (93 mg, m.p. 238°-240°C.).

EXAMPLE 31 Tablet--(Total weight 359 mg)

    ______________________________________                                        9-(2-Formyloxyethoxymethyl)guanine                                                                      100mg                                               Lactose                   200mg                                               Starch                    50mg                                                Polyvinylpyrrolidone      5mg                                                 Magnesium stearate        4mg                                                 ______________________________________                                    

EXAMPLE 32 Tablet--(Total weight 359 mg)

    ______________________________________                                        2-Amino-9-(2-formyloxyethoxymethyl)adenine                                                               100mg                                              Lactose                    200mg                                              Starch                     50mg                                               Polyvinylpyrrolidone       5mg                                                Magnesium stearate         4mg                                                ______________________________________                                    

The following compounds are also preferred:

    __________________________________________________________________________    Compound                     m.p.                                             __________________________________________________________________________    9-[2-(p-Fluorosulfonylbenzamido)ethoxymethyl]adenine                                                       201-202° C.                               9-(2-Bromoacetamidoethoxymethyl)adenine                                       hydrogen oxalate             132-133° C.                               9-[1-(2-Hydroxyethoxy)octyl]adenine                                                                        121-123° C.                               6-Dimethylamino-9-[1-(2-hydroxyethoxy)ethyl]purine                                                         86-88° C.                                 9-(2-Amino-1-methylethoxy)methyladenine dihydrochloride                                                    181-182° C. (eff)                         9-(2-Hydroxyethoxymethyl)-6-mercaptopurine                                                                 decomposes                                       9-(2-Hydroxypropoxy)methyladenine                                                                          164-167° C.                               9-(1,3-dibenzyloxy-2-propoxymethyl)adenine                                                                 120.5-122.5° C.                           9-(2-Sulfamoyloxyethoxymethyl)adenine.                                                                     172-173.5° C.                             9-(2-N-Carbobenzoxyphenylalanylamidoethoxymethyl)adenine                                                   208-210° C.                               9-(1,3-Dibenzyloxy-2-propoxymethyl)-6-mercaptopurine                          monohydrate                  162-164° C.                               9-(3-Benzoylpropoxymethyl)guanine                                                                          198-201° C.                               9-(2-Benzoyloxyethoxymethyl)purine                                                                         128-130° C.                               9-[1-(2-Hydroxyethoxy)ethyl]guanine                                                                        >260° C.                                  9-Ethoxymethylguanine        275-280° C. (dec.)                        9-[(2-Amino-1-cyclopentylethoxy)methyl]-                                      6-dimethylaminopurine dihydrochloride                                                                      153-154° C. (dec.)                        9-[(2-Amino-1-methylethoxy)methyl]-6-                                         dimethylaminopurine hydrochloride                                                                          201-203° C.                               9-[(2-N-Carbobenzoxyphenylalanylamido-1-                                      cyclopentylethoxy)methyl]-6-dimethylamino-                                    purine                       146-147° C.                               6-Diethylamino-9-[1-(2-benzoyloxyethoxy)ethyl]purine                                                       83-86° C.                                 2-Amino-6-dimethylamino-9-[1-(2-hy-                                           droxyethoxy)ethyl]purine hamlhydrate                                                                       92-94° C.                                 2-Amino-6-chloro-9-[1-(2-benzoyloxy-                                          ethoxy)ethyl]purine          125-130° C.                               2-Amino-6-chloro-9-(4-benzoyloxy-                                             butoxymethyl purine          119-121° C.                               9-[(2-N-Carbobenzoxyphenylalanylamido-1-                                      methylethoxy)methyl]-6-dimethylaminopurine                                                                 149-152° C.                               2-Amino-9-(2-benzoyloxyethoxymethyl)purine                                                                 149-154° C.                               9-(2-Benzoyloxyethoxymethyl)-2,6-di-                                          azidopurine                  124.5-125.0° C.                           9-Carboxymethoxymethyl-2,6-diamino-                                           purine . 0.02 2-methoxyethanol                                                                             ˜250° C. (dec.)                     9-(4-Hydroxy-n-butoxymethyl)guanine                                                                        234° C. with resolidifi-                  hemihydrate                  cation and decomposition                         6-Dimethylamino-9-(2-hydroxypropoxy)-                                         methylpurine hydrochloride   144-146° C.                               9-[(2-Phenylalanylamido-1-methylethoxy)                                       methyl]-6-dimethylaminopurine one                                             quarter hydrate              77-80° C.                                 9-[(2-Phenylalanylamido-1-phenylethoxy)                                       methyl]-6-dimethylaminopurine one                                                                          sinter ˜135° C.                     quarter hydrate              melt 142-144° C.                          2-Acetamido-9-(2-acetyloxyethoxymethyl)hypoxanthine                                                        202.5-204.5° C.                           __________________________________________________________________________

The above compounds were prepared by the methods disclosed herein.

EXAMPLE 34 Treatment of Herpes Simplex

Both eyes of a New Zealand White rabbit were infected with a suspensionof the PH8 strain of type I herpes simplex virus using the teaching ofthe method of Jones, B. J. Wise, J. B. and Patterson A. entitled Themeasurement of enhancement or inhibition of virus replication in thecornea. Evaluation of drug effects in the eye, 83- 97. Symposium of theRoyal Society of Medical, ed. Pigott, P. V., Association of MedicalAdvisers to the Pharmaceutical Industry, London, 1968.

One of the infected eyes of the rabbit was then treated topicallybeginning on the 4th day after infection with two drops of a 1% aqueoussolution of 2-amino-9-(2-hydroxyethoxymethyl) adenine for a period offour days 5 times per day. On the fifth day after treatment began thetreated infected eye was free of infection whereas the non-treated eyecontinued to exhibit the infection.

EXAMPLE 35 Treatment of Herpes Simplex

The method of Example 33 was followed except that a 1% aqueous solutionof 9-(2-hydroxymethyl) guanine was administered beginning on the 3rd dayafter infection. The results were the same as in Example 33.

EXAMPLE 36 9-(2-Benzoyloxyethoxymethyl)-2-methylthioadenine

To a solution of 6.00 g (28.4 mmol) of7-formamido-5-methylthiofurazano[3,4-d]pyrimidine¹ in 50 ml ofdimethylformamide which was cooled on an ice bath and protected frommoisture with a calcium chloride drying tube was added 3.00 g (29.7mmol) of triethylamine. After five minutes 8.05 g (37.6 mmol) of2-benzoyloxyethoxymethyl chloride was added along with 5 ml ofdimethylformamide. An additional 1.00 g (9.9 mmol) of triethylamine wasadded, the ice bath was removed, and the reaction was stirred at ambienttemperature overnight. The reaction was poured over 400 g of crushed icecontaining 2 ml of acetic acid. The mixture was extracted with five 50ml portions of chloroform, the combined extracts were washed with 25 mlof brine and dried (MgSO₄). The solvent was removed by spin evaporationin vacuo at asperator pressure and finally at about 1 mm Hg to removethe dimethylformamide. The resultant dark oil was dissolved in 200 ml ofethyl acetate and passed through a pad of superfiltrol #9. The pad waswashed with two 75 ml portions of ethyl acetate. The combined filtratesand washes were spin evaporated in vacuo to give an orange oil which wasa mixture (˜1 to 1) of formylated and unformylated product. The oil wastherefore dissolved in formic acetic anhydride (formed from 100 ml ofacetic anhydride and 50 ml of 97-100% formic acid) and heated on an oilbath at 80° for two hours, left overnight at ambient temperature andthen spin evaporated in vacuo to give7-(M-formyl-2-benzoyloxyethoxymethylamino)-5-methylthiofurazano[3,4-d]pyrimideas a crude oil. This oil was dissolved in 200 ml of acetic acid andplaced on a water bath. While the solution was magnetically stirred 30 gof zinc powder was added in portions over 20 minutes. The mixture wasthen heated near reflux for 1.5 hr, cooled, filtered, the solids washedwith acetic acid and then spin evaporated in vacuo. The residue wascovered with 100 ml of water and turned on the rotoevaporator withoutheating until a solid formed. The aqueous layer was decanted, the solidswere digested with 50 ml of ethanol and allowed to cool. The solids werecollected, washed with ether and dried; yield, 3.30 g (32%), m.p.166°-167°. Recrystallization of 0.80 g from ethanol gave the analyticalsamaple of 9-(2-benzoyloxyethoxymethyl)-2-methylthioadenine, m.p.166°-168°; λmax (ε×10⁻¹) (10% EtOH in solvent) pH 1-270 (15.0), 300 (sh)(7.0), H₂ O-276 (13.6), pH 13-276 (14.9) nm; nmr (DMSO-D₆), 2.50 (3H,S), 3.94 (2H, M), 4.40 (2H,m), 5.60 (2H, S), 7.33 (2H, S), 7.43-8.01(5H, M), 8.16 (1H, S)ε.

Anal. Calc'd. for C₁₆ H₁₇ N₅ O₃ S: C, 53.47; H, 4.77; N, 19.49. Found:C, 53.72; H, 4.85; N, 19.56.

EXAMPLE 37 9-(2-Hydroxyethoxymethyl)-2-methylthioadenine

A mixture of 2.20 g (6.12 mmol) of9-(2-benzoyloxyethoxymethyl)-2-methylthioadenine and 100 ml of 40%aqueous methylamine was heated on a steam bath with occasional agitationfor one hour. The solution was cooled and spin evaporated in vacuo. Theresultant oil was leached with five 40 ml portions of hot ether to givea solid; yield, 1.31 g (84%), m.p. 174°-176°. Recrystallization fromethanol gave the analytical sample of9-(2-hydroxyethoxymethyl)-2-methylthioadenine; λmax (ε×10⁻³) (10% EtOHin solvent) pH 1-270 (16.7),291 (sh) (11.9), H₂ O and pH 13-276 (15.2)nm; nmr (DMSO-d₆), 2.49 (3H, S), 3.55 (4H, S), 4.65 (1H, br S), 5.55(2H, S), 7.31 (2H, S), 8.15 (1H, S)ε.

Anal. Calc'd for C₉ H₁₃ N₅ O₂ S: C, 42.34; H, 5.13; N, 27.43. Found: C,42.66; H, 5.10; N, 27.39.

EXAMPLE 38 9-(2-Hydroxyethoxymethyl)-2-methylthiohypoxanthine

To a stirred solution of 0.50 g (1.96 mmol) of9-(2-hydroxyethoxymethyl)-2-methylthioadenine in 10 ml of AcOH was added0.83 g (12.0 mmol) of NaNO₂ in portions over a period of 1 hour. Thereaction mixture was stirred overnight, the white solids were removed byfiltration and washed with a few mls of AcOH. The combined filtrates andwashings were spin evaporated in vacuo to give a water soluble solidwhich was dissolved in 40 ml of H₂ O. This solution was applied to anAmberlite XAD-2 column [23 cm×3 cm] which was eluted with 1000 ml of H₂O to remove contaminating salts. Elution with 30% EtOH in H₂ O removedthe product which was isolated by spin evaporation and recrystallizationfrom EtOH; yield, 0.245 g (48%) of9-(2-hydroxyethoxymethyl)-2-methylthiohypoxanthine, m.p. 190°-193°, λmax(ε×10⁻³) (10% EtOH in solvent) pH 1-268 (16.2), H₂ O-262 (15.2), 284(sh) (11.5), pH 13-272 (15.2) nm; nmr (DMSO-d₆), 2.58 (3H,s), 3.57(4H,s), 4.67 (1H, br s), 5.56 (2H,s), 8.14 (1H,s), 12.52 (1H, br s) δ.

Anal. Calc'd. for C₉ H₁₂ N₄ O₃ S: C, 42.17; H, 4.72; N, 21.86. Found: C,42.36; H, 4.72; N, 21.57.

EXAMPLE 39 9-(2-Hydroxyethoxymethyl)guanine

A solution of 44 mg of9-(2-hydroxyethoxymethyl)-2-methylthiohypoxanthine in 50 ml of NH₃saturated EtOH was heated in a stainless steel vessel at 140° for 60 hr.The reaction solution was spin evaporated in vacuo and the residualsolid was recrystallized from EtOH; yield, 15 mg (39%). The tlc, UV,NMR, and mass spectral data were the same as that for authentic9-(2-hydroxyethoxymethyl)guanine prepared in Example 5.

EXAMPLE 40 9-(2-Acetyloxyethoxymethyl)guanine

A mixture of 9-(2-hydroxyethoxymethyl)guanine (4.6 g), drydimethylformamide (46 ml), acetic anhydride (16 ml) and dry pyridine (24ml) was stirred at room temperature overnight. The resulting white solidwas removed by filtration and dissolved in warm dimethylformamide (100ml), pyridine (10 ml) and acetic anhydride (8 ml) added and the mixturestirred for 18 hr. The white solid formed was removed by filtration,washed with ethyl acetate and recrystallized from dimethylformamide togive 9-(2-acetyloxyethoxymethyl)guanine (3.3 g), m.p. 240°-241° C.

EXAMPLE 41 9-(2-Propionyloxyethoxymethyl)guanine

A mixture of 9-(2-hydroxyethoxymethyl)guanine (1.0 g) and drydimethylformamide (50 ml) was heated on a steam bath until most of thesolid had dissolved. It was then cooled to room temperature. Drypyridine (10 ml) and propionic anhydride (2.9 ml) was added and themixture stirred at room temperature overnight. Additional propionicanhydride (1.0 ml) was added and the mixture stirred for an additional18 hr. The reaction mixture was diluted with ethyl acetate, chilled andthe resulting solid removed by filtration. This was recrystallized fromdimethylformamide to give 9-(2-propionyloxyethoxymethyl)guanine (0.9 g),m.p. 233°-226° C.

EXAMPLE 42 9-[2-(2,2-Dimethylpropionyloxy)ethoxymethyl]guanine

A mixture of 9-(2-hydroxyethoxymethyl)guanine (2.46 g), dry pyridine(400 ml), and pivalic anhydride (6.5 ml) was heated on a steam bath fora total of 33 days. On day 11 additional pyridine (150 ml) was added,and on day 27 dimethylformamide (50 ml) was added. Volatiles wereremoved under reduced pressure, and the residue was triturated withethyl acetate. The insoluble solid was removed by filtration anddissolved in methanol-acetone. Silica gel (3 g) was added and thesolvent evaporated. The residue was added to a column of silica gel (180g) in acetone. Elution with acetone yielded an initial fraction ofN,O-diacylated material followed by a fraction containing the desiredmonoacylated product. The acetone was evaporated from this latterfraction, and the residue was recrystallized fromdimethylformamide-acetonitrile-ethyl acetate to give9-[2-(2,2-dimethylpropionyloxy)ethoxymethyl]guanine (0.5 g), m.p.245°-246° C.

EXAMPLE 43 9-(2-Aminoethoxymethyl)guanine

A dispersion of N-(2-hydroxyethyl)phthalimide (19.1 g) andparaformaldehyde (3.0 g) in 1,2-dichloroethane (250 ml) was cooled in anice-salt-acetone bath and saturated with dry hydrochloric acid withstirring. After 4 hours the mixture was dried over calcium chloride,filtered and evaporated under reduced pressure to giveN-(2-chloromethoxyethyl)phthalimide (21.9 g), m.p. 69°-72° C. To astirred solution of tris trimethylsilylguanine (16.5 g) in toluene (50ml) was added N-(2-chloromethoxyethyl)phthalimide (17.0 g) andtriethylamine (23 ml). The reaction mixture was heated at reflux undernitrogen for 29 hours, cooled and evaporated under reduced pressuregiving a dark brown oil. The oil was digested in ethanol (400 ml) on asteam bath for 40 minutes, giving a solid which was collected and washedwith ethanol and ether. It was recrystallized from 2-methoxyethanol togive 9-(2-phthalimidoethoxymethyl)guanine hydrate (14.4 g), m.p.>240° C.(dec.). A mixture of 9-(2-phthalimidoethoxymethyl)guanine hydrate (0.94g), hydrazine (2 ml) and ethanol (100 ml) was heated at reflux withstirring for one hour. The reaction mixture was filtered hot, and thefiltrate was cooled and evaporated under reduced pressure. Ethanol wasadded and the mixture re-evaporated. This was repeated three times. Theresidual solid was stirred with 3% aqueous acetic acid (50 ml) for 30minutes and filtered. The solids were washed with water (15 ml), and thewash and filtrate combined. The combined wash and filtrate wasevaporated under reduced pressure at<35° C. to a small volume, dilutedwith ethanol and evaporated to dryness. The pasty, white solid wasdispersed in a small amount of ethanol and diluted with a few ml etherto give a granular solid which was collected, washed with ethanol anddried. Recrystallization from ethanol-water gave9-(2-aminoethoxymethyl)guanine acetate monohydrate (0.43 g), m.p. (˜135°C. melts and resolidifies) 174°-176° C.

Example 44 9-(2-hydroxyethoxymethyl)guanine

To a mixture of acetic anhydride (1.2 ml) and p-toluenesulfonic acid(0.09 g) was added with stirring dioxolane (1.02 g)-caution, exothermic.The solution was allowed to cool for several minutes. Diacetylguanine(1.45 g) and dry toluene (9 ml) were added and the reaction mixture wasstirred at reflux for 18 hr. and then allowed to cool to roomtemperature. The toluene was decanted off and the residue trituratedseveral times with benzene. Methanol (10 ml) was added to the residueand evaporated under reduced pressure. To the residue was added 40%aqueous methylamine (10 ml) and the mixture heated on a steam bath for20 min. The water and methylamine were removed under reduced pressure,ethanol (10 ml) was added and evaporated. The residue was thoroughlyextracted with boiling methanol (700 ml total) and the combined extractsevaporated. The resulting solid was triturated with cold ethanol andthen recrystallized from methanol to give9-(2-hydroxyethoxymethyl)guanine (0.3 g).

I claim:
 1. A compound of the formula (I) ##STR12## wherein R¹ is amino,R² is aminoX is oxygen or sulphur R³ is hydrogen, straight or branchedchain or cyclic alkyl, hydroxyalkyl, benzyloxyalkyl or phenyl; R⁴ ishydrogen, hydroxy or alkyl; R⁵ is hydroxy, amino, alkyl, hydroxyalkyl,benzyloxy, benzoyloxymethyl, sulphamoyloxy, phosphate, or acyloxy having1 to 8 carbon atoms, R⁶ is hydrogen or alkyl; or a pharmaceuticallyacceptable salt thereof.
 2. A pharmaceutical composition for use as anantiviral which comprises an effective antiviral amount of the compoundor salt of claim
 1. 3. A method of treating a susceptible virusinfection in a mammal which comprises the administration to the infectedmammal of an effective, non-toxic antiviral amount of the compound orsalt of claim
 1. 4. The compound or salt of claim 1 in which X isoxygen.
 5. The composition of claim 2 in which X is oxygen.
 6. Themethod of claim 3 in which X is oxygen.
 7. The compound or salt of claim1 in which R⁵ is acetoxy.
 8. The method of claim 3 in which the virusinfection is a herpes virus infection.
 9. The compound or salt of claim1 in which the salt is an acid addition salt.
 10. The compound or saltof claim 1 in which R⁵ is hydroxy or phosphate.
 11. The composition ofclaim 2 in which R⁵ is hydroxy or phosphate.
 12. The method of claim 3in which R⁵ is hydroxy or phosphate. 13.2-amino-9-(2-hydroxyethoxymethyl)adenine.
 14. A pharmaceuticallyacceptable salt of 2-amino-9-(2-hydroxyethoxymethyl)adenine.
 15. Thesalt of claim 14 in which the salt is an acid addition salt.
 16. Themethod of treating a susceptible viral infection in a mammal having saidviral infection which comprises the administration to said mammal of aneffective antiviral non-toxic amount of2-amino-9-(2-hydroxyethoxymethyl)adenine or a pharmaceuticallyacceptable salt thereof.
 17. The method of claim 16 in which theadministration is oral, parenteral or topical.
 18. The method of claim16 in which the viral infection is a herpes virus infection.
 19. Apharmaceutical composition comprising an effective antiviral amount of2-amino-9-(2-hydroxyethoxymethyl)adenine or a pharmaceuticallyacceptable salt thereof together with a pharmaceutically acceptablecarrier therefore.
 20. A pharmaceutical composition of claim 19 in aform for oral, parenteral or topical administration.
 21. Apharmaceutical composition for treating a susceptible viral infectioncomprising an effective antiviral treatment amount of2-amino-9(2-hydroxyethoxymethyl)adenine together with a pharmacueticallyacceptable carrier therefore.
 22. A method of treating a herpes virusinfection in a human which comprises administering to said human aneffective anti herpes treatment amount of the compound2-amino-9-(2-hydroxyethoxymethyl)adenine.
 23. The method of claim 22 inwhich the compound is administered in a pharmacuetically acceptablecarrier.
 24. The method of claim 18 in which the viral infection is aherpes simplex or herpes zoster infection.
 25. The method of claim 22 inwhich the herpes virus infection is a herpes simplex or herpes zosterinfection.